A Novel SPAST Mutation Results in Spastin Accumulation and Defects in Microtubule Dynamics

Abstract

Background Haploinsufficiency is widely accepted as the pathogenic mechanism of spastic paraplegia type 4 (SPG4). However, there are some cases that cannot be explained by reduced function of the spastin protein encoded by SPAST . Objectives To identify the causative gene of autosomal dominant hereditary spastic paraplegia in three large Chinese families and explore the pathological mechanism of a spastin variant. Methods Three large Chinese hereditary spastic paraplegia families with a total of 247 individuals (67 patients) were investigated, of whom 59 members were recruited to the study. Genetic testing was performed to identify the causative gene. Western blotting and immunofluorescence were used to analyze the effects of the mutant proteins in vitro. Results In the three hereditary spastic paraplegia families, of whom three index cases were misdiagnosed as other types of neurological diseases, a novel c.985dupA (p.Met329Asnfs*3) variant in SPAST was identified and was shown to cosegregate with the phenotype in the three families. The c.985dupA mutation produced two truncated mutants (mutant M1 and M87 isoforms) that accumulated to a higher level than their wild‐type counterparts. Furthermore, the mutant M1 isoform heavily decorated the microtubules and rendered them resistant to depolymerization. In contrast, the mutant M87 isoform was diffusely localized in both the nucleus and the cytoplasm, could not decorate microtubules, and was not able to promote microtubule disassembly. Conclusions SPAST mutations leading to premature stop codons do not always act through haploinsufficiency. The truncated spastin may damage the corticospinal tracts through an isoform‐specific toxic effect.

A novel truncating variant of SPAST associated with hereditary spastic paraplegia indicates a haploinsufficiency pathogenic mechanism

“…To date, different mechanistic hypotheses for the etiology of truncating variants in SPAST-HSP have been proposed. We have summarized the pathogenic truncating variants that have had transcription or protein expression analysis performed in previous studies (Supplementary Table 4) (16, [18][19][20][21][22][23][24][25][26]. Haploinsufficiency has been the explanation for truncating SPAST mutations in the majority of the literature.…”

“…However, some truncating variants (c.550dupT, c.734C > G, and c.985dupA) escape degradation and are stable. Their truncated mutant proteins could accumulate to a higher level than their wild-type counterparts, indicating possible gain-of-function mechanisms ( 18 , 20 ). The authors concluded that premature stop codons caused by SPAST mutations do not always result in haploinsufficiency ( 18 ).…”

“…Their truncated mutant proteins could accumulate to a higher level than their wild-type counterparts, indicating possible gain-of-function mechanisms ( 18 , 20 ). The authors concluded that premature stop codons caused by SPAST mutations do not always result in haploinsufficiency ( 18 ). They hypothesized that the position of the premature termination codon from an exon-exon junction in SPAST may be crucial to preventing nonsense-mediated mRNA decay.…”

“…They hypothesized that the position of the premature termination codon from an exon-exon junction in SPAST may be crucial to preventing nonsense-mediated mRNA decay. Moreover, the less stable mRNAs with premature termination codons might be translated into truncated spastin proteins with greater stability ( 18 , 20 ). Therefore, it would be important to perform transcription and protein expression analysis for different truncating SPAST pathogenic variants, as these studies would provide insight into the mechanisms underlying their pathogenicity.…”

The Puzzle of Hereditary Spastic Paraplegia: From Epidemiology to Treatment

“…Microtubules encourage the growth of axons in central nervous system neurons and help maintain the neurons’ morphology, transport, and motility, along with maintaining the complexity of neutrine arrangement and degradation. Spastin protein encoded by the SPAST gene is necessary for maintaining microtubule dynamics, and its mutation is linked with SPG4 [ 37 ]. Spastin serves microtubules in more stable regions by targeting tubulin post-translation [ 38 , 39 ].…”